Testing the sample quality Flashcards
What is genomics used for?
Ancestry
Crime
Disease
What event revolutionised genomics?
Human Genome Project
2003
What is the most important part of a scientific experiment?
Obtaining an appropriate sample
What characterises as an appropriate sample in microarrays?
Good quality
What are ways in which the source used in genome investigations can differ?
Source
Type
Species
Different sources of genetic samples
Cells
Tissues
Blood
Saliva
Bacteria
Different types of genetic samples
Fresh
Cultured
Frozen
Fixed
Archived
Laser capture
Different species of genetic samples
Human
Mouse
Fish
Insect
What 3 things must we know about a sample in order to carry out an investigation?
Concentration
Integrity
Contaminants
What does agarose gel help us determine?
Whether our DNA sample is degraded or not
How does agarose gel tell us whether DNA is degraded or not?
Nucleic acid, which is negatively charged, will move towards the positive electrode and form bands
The less concentrated the DNA sample is at the electrode end, the lower quality the sample is since it is more degraded
What is the limitation of agarose gel?
Qualitative
Not tell the investigator about the quality or concentration of the sample
Where should a high quality DNA give a major band on the agarose gel?
10-20kb
What is nanodrop useful for?
Assessing the purity of DNA and RNA
How big are the samples measured by a nandrop?
1 nanol
Describe how a nanodrop assesses the purity of DNA or RNA
Measures the absorbance of the sample at 230, 260 and 280
DNA and RNA absorb wavelengths at 260
230 = salt contamination 280 = protein contamination
What are the results of a sample?
Ratio of the 260/230 and 260/280 absorbances
What is the normal ratio of a pure DNA sample?
1.8-2.0
What is the normal ratio of a pure RNA sample?
2.0-2.2
How can absorbance be used to measure the sample concentration?
Concentration = (absorbance X (e = 50 (DNA) and 40(RNA)) / path length in cm
What does a low ratio mean?
High concentration of salt/protein compared to nucleic acid
What is the shape of the absorbance curve of a good sample?
Peak at 260
Does not start at origin
What is the shape of the absorbance curve of a bad sample?
Peak at 230/280 depending on what the contaminant is
If a sample is contaminated, that automatically means we cannot use it
TRUE or FALSE
FALSE
We need to clean the sample using a mesh to remove the contaminants